Signal transduction pathways control cell decisions and cell fate, and when not functioning properly can give rise to diseases such as cancer. The yeast pheromone response pathway is a model signal transduction pathway that regulates transcription, cell cycle, and cell morphology, and can be attenuated should mating not occur. Many of the factors involved in this pathway remain to be discovered. For example, the Rgalp protein was recently identified by using a selection for constitutive activators (negative regulators) of the pathway. Rgalp was shown to be a negative regulator of the pheromone response pathway that has homology to mammalian GAP and LIM domains. Two hybrid analysis demonstrated that Rgalp interacts with the polarity establishment protein Cdc42p, presumably to affect changes in cell morphology during mating. This proposal aims to identify and characterize negative regulators of the pheromone response pathway. In particular, recessive constitutive mutants of the pathway that were identified in the same selection that uncovered RGA1 will be cloned and characterized genetically. In addition, two new mutant isolation selections will be used to isolate novel negative regulators of the pathway. Characterization of these mutants will lead to a more complete understanding of eukaryotic signal transduction.